Gut microbiota carcinogen metabolism causes distal tissue tumours.

Exposure to environmental pollutants and human microbiome composition are important predisposition factors for tumour development1,2. Similar to drug molecules, pollutants are typically metabolized in the body, which can change their carcinogenic potential and affect tissue distribution through altered toxicokinetics3. Although recent studies demonstrated that human-associated microorganisms can chemically convert a wide range of xenobiotics and influence the profile and tissue exposure of resulting metabolites4,5, the effect of microbial biotransformation on chemical-induced tumour development remains unclear. Here we show that the depletion of the gut microbiota affects the toxicokinetics of nitrosamines, which markedly reduces the development and severity of nitrosamine-induced urinary bladder cancer in mice6,7. We causally linked this carcinogen biotransformation to specific gut bacterial isolates in vitro and in vivo using individualized bacterial culture collections and gnotobiotic mouse models, respectively. We tested gut communities from different human donors to demonstrate that microbial carcinogen metabolism varies between individuals and we showed that this metabolic activity applies to structurally related nitrosamine carcinogens. Altogether, these results indicate that gut microbiota carcinogen metabolism may be a contributing factor for chemical-induced carcinogenesis, which could open avenues to target the microbiome for improved predisposition risk assessment and prevention of cancer.

MyD88 Signaling Accompanied by Microbiota Changes Supports Urinary Bladder Carcinogenesis.

Urinary bladder cancer (BC) inflicts a significant impairment of life quality and poses a high mortality risk. Schistosoma haematobium infection can cause BC, and the urinary microbiota of BC patients differs from healthy controls. Importantly, intravesical instillation of the bacterium Bacillus Calmette-Guerin stands as the foremost therapy for non-muscle invasive BC. Hence, studying the receptors and signaling molecules orchestrating bacterial recognition and the cellular response in the context of BC is of paramount importance. Thus, we challenged Toll-like receptor 4 (Tlr4) and myeloid differentiation factor 88 (Myd88) knock-out (KO) mice with N-butyl-N-(4-hydroxylbutyl)-nitrosamine (BBN), a well-known urinary bladder carcinogen. Gut microbiota, gene expression, and urinary bladder pathology were followed. Acute exposure to BBN did not reveal a difference in bladder pathology despite differences in the animal's ability to recognize and react to bacteria. However, chronic treatment resulted in reduced cancer invasiveness among Myd88KO mice while the absence of functional Tlr4 did not influence BC development or progression. These differences correlate with a heightened abundance of the Faecalibaculum genus and the lowest microbial diversity observed among Myd88KO mice. The presented data underscore the important role of microbiota composition and MyD88-mediated signaling during bladder carcinogenesis.

The dynamics of the inflammatory response during BBN-induced bladder carcinogenesis in mice.

BACKGROUND: Bladder cancer (BC) is the most common malignant disease of the urinary tract. Recurrent high grade non muscle invasive BC carries a serious risk for progression and subsequent metastases. The most common preclinical mouse model for bladder cancer relies on administration of N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) to mice. BBN-induced tumors in mice recapitulate the histology of human BC and were characterized with an overexpression of markers typical for basal-like cancer subtype in addition to a high mutational burden with frequent mutations in Trp53, similar to human muscle invasive BC. METHODS: Bladder cancer was induced in C57BL/6J male mice by administering the BBN in the drinking water. A thorough histopathological analysis of bladder specimen during and post BBN treatment was performed at 2, 4, 16, 20 and 25 weeks. RNA sequencing and qPCR was performed to assess the levels of expression of immunologically relevant genes at 2 weeks and 20 weeks during and post BBN treatment. RESULTS: We characterized the dynamics of the inflammatory response in the BBN-induced BC in mice. The treatment with BBN had gradually induced a robust inflammation in the first 2 weeks of administration, however, the inflammatory response was progressively silenced in the following weeks of the treatment, until the progression of the primary carcinoma. Tumors at 20 weeks were characterized with a marked upregulation of IL18 when compared to premalignant inflammatory response at 2 weeks. In accordance with this, we observed an increase in expression of IFNγ-responsive genes coupled to a pronounced lymphocytic infiltrate during the early stages of malignant transformation in bladder. Similar to human basal-like BC, BBN-induced murine tumors displayed an upregulated expression of immunoinhibitory molecules such as CTLA-4, PD-L1, and IDO1 which can lead to cytotoxic resistance and tumor escape. CONCLUSIONS: Despite the recent advances in bladder cancer therapy which include the use of checkpoint inhibitors, the treatment options for patients with locally advanced and metastatic BC remain limited. BBN-induced BC in mice displays an immunological profile which shares similarities with human MIBC thus representing an optimal model for preclinical studies on immunomodulation in management of BC.

Expression of Mitochondrial Respiratory Chain Complexes in the Vaginal Wall in Postmenopausal Women with Pelvic Organ Prolapse.

BACKGROUND/AIMS: Pelvic organ prolapse (POP) is a common disease affecting adult women. It is a result of the vaginal wall disorder as well as damage of the supportive structures contributing to the integrity of the pelvic floor. Mitochondrial disorders may have an important role in the vaginal wall degeneration leading to POP. The goal of this research is to examine if POP is associated with an altered expression of mitochondrial respiratory chain complexes. METHODS: Samples of vaginal tissue were collected from 16 postmenopausal women: 10 had POP and 6 had other forms of benign gynecological disease. Using western blot, samples were analyzed to assess the expression of mitochondrial proteins including citrate synthase (CS), individual complexes of the mitochondrial respiratory chain and alpha smooth muscle actin (SMA). RESULTS: A significantly reduced expression of SMA and complex II in vaginal tissue of women with POP was found, compared to the control group (p < 0.05), with a tendency for a reduced expression of CS (p = 0.06) and other complexes in the POP group. CONCLUSIONS: Our results indicate that there is a decreased quantity of the smooth muscle and a decreased expression of mitochondrial markers in the vaginal wall of women with prolapse suggesting their possible role in the pathogenesis of POP.

Lateral thermal damage of mesoappendix and appendiceal base during laparoscopic appendectomy in children: comparison of the harmonic scalpel (Ultracision), bipolar coagulation (LigaSure), and thermal fusion technology (MiSeal).

BACKGROUND: The aim of this study was to compare lateral thermal damage of mesoappendix and appendiceal base using three different instruments for sealing and cutting of mesoappendix. MATERIALS AND METHODS: A total number of 99 patients (54 males and 45 females) who underwent laparoscopic appendectomy because of suspected appendicitis between December 2013 and May 2015 were enrolled in the study. The patients were divided in three groups based on instrument used for sealing of mesoappendix: group 1 (Ultracision; n = 36), group 2 (LigaSure; n = 32), and group 3 (MiSeal; n = 31). Lateral thermal damage, intraoperative and postoperative complications, duration of surgery, hospital stay, and economic value were compared within groups. RESULTS: The median age of patients was 14 y (range 3-17). A histopathologic analysis revealed a positive diagnosis of appendicitis in 84 patients (85%). The median lateral thermal damage on appendiceal base using Ultracision, LigaSure, and MiSeal was 0.10 mm, 0.16 mm, and 0.10 mm respectively, and on mesoappendix, 0.08 mm, 0.13 mm, and 0.08 mm, respectively. Significantly higher thermal damage was found on mesoappendix (P = 0.015) and appendiceal base (P = 0.012) in patients treated with LigaSure than in patients from other groups. There were no statistical differences among the groups regarding intraoperative and postoperative complications (P = 0.098). No significant difference in thermal damage between appendicitis and nonappendicitis group was found (P = 0.266). CONCLUSIONS: Using of Ultracision, LigaSure, and MiSeal for sealing of mesoappendix in laparoscopic appendectomy in children is safe and useful. LigaSure produces significantly greater lateral thermal damage compared with other instruments.

Immunohistochemical expression of hypoxia-inducible factor-1α in stromal cells of vaginal tissue in post-menopausal women with pelvic organ prolapse.

Background & objectives: Pelvic organ prolapse (POP) is a common medical condition that affects adult women of different ages. The support of a normal pelvic floor is the result of complex interactions between ligaments, muscles, connective tissue and vaginal walls. Hypoxia and oxidative stress can reduce protein synthesis in the pelvic muscles that may contribute to muscular atrophy. Hypoxia-inducible factor-1α (HIF-1α) is a transcriptional activator which, expressed in response to hypoxia, activates a number of genes involved in cellular response to hypoxia. However, a potential role of hypoxia and oxidative stress in pathogenesis of POP is not known. This study was aimed to compare the level of HIF-1α immunohistochemical expression in the vaginal stromal cells of postmenopausal women with and without POP. Methods: Samples of the vaginal tissue from 120 menopausal women were obtained during surgery, and immunohistochemical expression of HIF-1α was assessed. There were 60 women with POP while 60 women in the control group were without prolapse but with benign gynaecological diseases. Results: In post-menopausal women with prolapse, significant differences were observed in the number of HIF-1α-positive stromal cells in the vaginal tissue compared to the control group. There was a significant increase in the number of HIF-1α in the stromal cells of the vaginal tissue in women with prolapse. Interpretation & conclusions: Difference in expression of HIF-1α in stromal cells of the vaginal tissue in the post-menopausal women with and without POP suggests that prolonged hypoxia probably has an important role in the aetiopathogenesis of POP.

Protective effect of urapidil on testicular torsion-detorsion injury in rats.

PURPOSE: The aim of this study was to investigate the effect of urapidil and low-molecular weight heparin (LMWH) on testicular torsion-detorsion injury in rats. METHODS: Thirty-two male Sprague-Dawley rats were used. In the torsion-detorsion (T/D) group, the left testis was twisted at 720° for 3 h. After 3 h of reperfusion, the testis was removed. Urapidil or LMWH was administered intraperitoneally 30 min before detorsion in the treatment groups. RESULTS: Unilateral testicular torsion-detorsion caused significant increases in the malondialdehyde level and apoptosis and significant decreases in the superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities in ipsilateral testes (p < 0.001). The rats treated with urapidil had a significant decrease in the malondialdehyde level and apoptosis and significant increases in the SOD and GPx activities in ipsilateral testes compared to the T/D group (p < 0.001). Animals treated with LMWH showed non-significant reductions in malondialdehyde levels and apoptosis compared to the T/D group. In addition, no significant difference in the SOD activities (p = 0.52) between the groups was found. The increase in the GPx activities was significant in the LMWH group compared to the T/D group (p < 0.001). CONCLUSION: The administration of urapidil before detorsion prevents ischemia/reperfusion cellular damage in testicular tissue. LMWH was not found to have a beneficial effect on testicular T/D injury in rats.

Unusual cause of acute abdomen in a child--torsion of greater omentum: report of two cases.

INTRODUCTION: Torsion of the omentum is twisting along its long axis and a rare cause of acute abdomen. Depending on associated conditions, it is classified as primary and secondary. It may mimic different pathologies presenting as acute abdomen, most common of them being acute appendicitis. Current choice for management of omental torsion is laparoscopic surgery. CASE PRESENTATION: We present two cases of omental torsion of two boys who presented with abdominal pain, nausea and vomiting and underwent emergency laparoscopy. CONCLUSION: Omental torsion is very rare, and its diagnosis is usually made only after surgery. At laparoscopy, omental torsion is suspected when the appendix is normal and the symptoms and findings of torsion are present. Laparoscopy is a safe and effective approach for the diagnosis and management of omental torsion, with the advantages of reduced postoperative pain and hospital stay.

Which theory for the origin of syphilis is true?

INTRODUCTION: There are four theories about the origin of syphilis, of which the mostly represented one is the Columbian theory. This theory suggests that syphilis was brought into Europe in 1493 ad by the ship from Caribbean islands. AIM: The aim of this study is to test all theories on a sample of 403 skeletons: 135 from prehistory, 134 from antique, and 134 from medieval period and new age from the Dalmatia (Croatia). METHODS: All skeletons were examined using standard anthropological methods. Paleopathological analysis was performed on each skeleton as well as additional radiographic method on one isolated skeleton. MAIN OUTCOME MEASURES: Paleopathological changes on skeletal remains connected with treponematosis. RESULTS: Paleopathological analysis revealed one skeleton from the antique period (second to 6th century A.D.) that exhibited skeletal markers similar to those described in one clinical case in which congenital syphilis was confirmed by a Wasserman reaction. Skeletal remains of this person were examined macroscopically and radiographically, and the differential diagnostics eliminated other considered pathologies as well as trauma. CONCLUSIONS: The finding of skeletal markers of syphilis on a skeleton from the antique supports the theory of pre-Columbian syphilis origin.

The Impact of Phenotype of Inflammatory Bowel Diseases, Inflammation Activity and Therapy on Mucosal Mature Cd83+ Dendritic Cell.

Background: Crohn's disease (CD) and ulcerative colitis (UC) are well-defined phenotypes of chronic inflammatory bowel diseases (IBDs). A mechanism of inflammation in these diseases is partially controlled by the intestinal dendritic cell (DC). In this study, we observed a mature CD83+ DC in colonic bioptic samples, and its correlation with disease phenotype and activity. Methods: The study included 219 subjects: 100 with UC, 44 with CD and 75 healthy subjects. Colonic biopsy specimens were incubated with the primary antibody Anti-CD83. Intraepithelial CD83+ DCs were counted per 100 enterocytes. The presence of CD83+ DC was analysed according to the type of IBD, histopathologic inflammation activity and treatment outcome. Results: The presence of mature CD83+ DCs (0, ≥1) differed according to disease types of IBD (p = 0.001), histologic inflammation activity (p = 0.049) and applied therapy (p = 0.001). The odds for CD83+ DC presence were 5.2 times higher in the CD group than in the control/UC group. The odds for CD83+ DC presence were 2.6 times higher in subjects without inflammation or chronic inflammation than with acute inflammation. They were also 3.7 times higher in subjects without therapy. The cut-off value 0.5 CD83+ DC (Rock analysis area = 0.699; SE 0.046; p < 0.001; 95% CI: 0.609-0.788) had been assessed as a differentiation marker between UC and CD. Conclusion: Presence of CD83+ DC could be used as a possible parameter in distinction between UC and CD, as well as a predictor of inflammation activity and treatment outcome.

Subbrachial approach to humeral shaft fractures: new surgical technique and retrospective case series study.

BACKGROUND: There are few surgical approaches for treating humeral shaft fractures. Here we present our results using a subbrachial approach. METHODS: We conducted a retrospective case series involving patients who had surgery for a humeral shaft fracture between January 1994 and January 2008. We divided patients into 4 groups based on the surgical approach (anterior, anterolateral, posterior, subbrachial). In all patients, an AO 4.5 mm dynamic compression plate was used. RESULTS: During our study period, 280 patients aged 30-36 years underwent surgery for a humeral shaft fracture. The average duration of surgery was shortest using the subbrachial approach (40 min). The average loss of muscle strength was 40% for the anterolateral, 48% for the posterior, 42% for the anterior and 20% for the subbrachial approaches. The average loss of tension in the brachialis muscle after 4 months was 61% for the anterolateral, 48% for the anterior and 11% for the subbrachial approaches. Sixteen patients in the anterolateral and anterior groups and 6 patients in the posterior group experienced intraoperative lesions of the radial nerve. No postoperative complications were observed in the subbrachial group. CONCLUSION: The subbrachial approach is practical and effective. The average duration of the surgery is shortened by half, loss of the muscle strength is minimal, and patients can resume everyday activities within 4 months. No patients in the subbrachial group experienced injuries to the radial or musculocutaneous nerves.

Sex determination in skeletal remains from the medieval Eastern Adriatic coast - discriminant function analysis of humeri.

AIM: To investigate the usefulness of humerus measurement for sex determination in a sample of medieval skeletons from the Eastern Adriatic Coast. Additional aim was to compare the results with contemporary female population. METHODS: Five humerus measurements (maximum length, epicondylar width, maximum vertical diameter of the head, maximum and minimum diameter of the humerus at midshaft) for 80 male and 35 female medieval and 19 female contemporary humeri were recorded. Only sufficiently preserved skeletons and those with no obvious pathological or traumatic changes that could affect the measurements were included. For ten samples, analysis of DNA was performed in order to determine sex using amelogenin. RESULTS: The initial comparison of men and women indicated significant differences in all five measures (P<0.001). Discriminant function for sex determination indicated that as much as 85% of cases could be properly categorized, with better results in men (86%) than women (80%). Furthermore, the comparison of the medieval and contemporary women did not show significant difference in any of the measured features. Sex results obtained by anthropological and DNA analysis matched in all 10 cases. CONCLUSION: The results indicate that humerus measurement in Croatian medieval population may be sufficient to determine the sex of the skeleton. Furthermore, it seems that secular changes have not substantially affected contemporary population, suggesting that the results of this study are transferable to contemporary population as well.

[Clinical guidelines for diagnosing, treatment and monitoring patients with bladder cancer--Croatian Oncology Society and Croatian Urology Society, Croatian Medical Association]. / Klinicke upute za dijagnostiku, lijecenje i pracenje bolesnika oboljelih od raka mokracnog mjehura Hrvatskoga Onkolokog Drustva i Hrvatskoga Uroloskog Drustva Hrvatskoga Lijecnickog Zbora.

Urothelial cancer is the most common bladder cancer. Hematuria is the most common presenting symptom in patients with bladder cancer. The most common diagnostics of bladder cancer is performed by transurethral resection of bladder after which pathohistological diagnosis is set. It is necessary to determine whether the cancer penetrated in muscle layer (muscle-invasive cancer) or not (muscle-noninvasive cancer). Decision on therapeutic modality depends on the clinical stage of disease and on prognostic and risk factors. For muscle non-invasive bladder cancer transurethral resection is preferred with or without intravesical instillation of Bacillus Calmette-Guérin (BCG). For invasive cancer the method of choice is radical cystectomy. Radiotherapy is used in radical and palliative purposes. Metastatic disease is most frequently treated by chemotherapy metotrexate/vinblastine/doxorubicine/cisplatin (MVAC) or gemcitabine/cisplatin (GC). The purpose of this article is to present clinical recommendations to set standards of procedures and criteria in diagnostics, treatment and follow up of patients with bladder cancer in the Republic of Croatia.

[Clinical guidelines for diagnosing, treatment and monitoring patients with prostate cancer--Croatian Oncology Society and Croatian Urology Society, Croatian Medical Association]. / Klinicke upute za dijagnostiku, lijecenje i pracenje bolesnika oboljelih od raka prostate Hrvatskoga Onkoloskog Drustva i Hrvatskoga Uroloskog Drustva Hrvatskoga Lijecnickog Zbora.

Prostate adenocarcinoma is the second most common solid neoplasm in male population in Croatia. It rarely causes symptoms unless it is advanced. The finding of PSA rise is the most common reason for diagnostic workout. Treatment plan is based on TNM classification, Gleason score and PSA. Clinically localized disease is successfully treated by radical prostatectomy or radiotherapy with or without hormonal therapy. Locally advanced disease is treated with radiotherapy and hormonal therapy. Metastatic disease can be controlled for many years by androgen deprivation. For castration resistant disease appropriate treatment is chemotherapy or secondary hormonal therapy. The following paper presents the clinical guidelines in order to standardize procedures and criteria for the diagnosis, management, management, treatment and monitoring of patients with prostate cancer in the Republic of Croatia.

Treatment of Inflammatory Dentigerous Cyst Using a Surgical T Drain in a Child.

Dentigerous cysts are rarely reported in young children. They are usually asymptomatic and only identified when becoming significantly large. Treatment by enucleation may damage structures like the inferior alveolar nerve, maxillary sinus, or permanent teeth, thus reducing the child's quality of life. Therefore, conservative surgical treatment such as decompression is indicated. This case report describes the treatment and subsequent complete regression of an inflammatory dentigerous cyst based on the decompression method using a customized surgical tube in a 10-year-old girl. The innervation was preserved, and permanent teeth erupted.

Elevated Tumor Cell-Intrinsic STING Expression in Advanced Laryngeal Cancer.

Laryngeal cancer is the second most common malignancy of the head and neck, worldwide. Immunotherapy targeting checkpoint inhibitors has been approved for the treatment of patients with recurrent or metastatic laryngeal cancer but has a relatively low response rate and outcomes that leave many patients underserved. Targeting the cGAS-STING signaling pathway can potentially improve the activation of immune effector cells, although its role in the development and progression of laryngeal cancer has not yet been investigated in depth. Fifty-nine tumor samples from patients with pathologically confirmed squamous cell carcinoma of the larynx, stage I-IV non-metastatic disease, who were treated at the University Hospital of Split, were immunohistochemically stained for the expression of STING, cGAS, CD8, CD68, and CD163. Elevated tumor cell-intrinsic STING expression was positively associated with stage IV (p = 0.0031), pT3, and pT4 laryngeal cancers (p = 0.0336) as well as with higher histological grades (G2 and G3) (p = 0.0204) and lymph node-positive tumors (p = 0.0371). After adjusting for age, sex, location, and cGAS expression, elevated STING expression was significantly associated with stage IV cancer in a multiple logistic regression model (ß = 1.849, SE = ±0.8643, p = 0.0324). Elevated STING expression represents a potentially favorable predictive biomarker for new therapeutic approaches involving STING agonists combined with immunotherapy and DNA-damaging agents (radiotherapy, cisplatin, and PARP inhibitors) in laryngeal cancer.

Her-2/neu assessment for gastric carcinoma: validation of scoring system.

BACKGROUND/AIMS: Gastric cancer is the second leading cause of cancer mortality in the world. Amplification of HER-2/neu oncogene has become an important biomarker for identifying patients who respond to HER-2 targeting therapy. A number of studies have analyzed HER-2/neu overexpression in gastric carcinoma, and the rate of HER2 positivity is variable, ranging from 6% to 35%. METHODOLOGY: In our study HER-2/neu expression was assessed on 73 samples of primary gastric cancer, using immunohistochemistry. For 19 patients preoperative biopsy samples and resected specimens were available. Additionally, internal ring study was performed to estimate intraobserver variability of IHC scoring among pathologists at our department. RESULTS: HER-2/neu overexpression was found in 10 (13.6%) of the tested samples, and it was more common in intestinal (22.5%) than the diffuse type (3.7%). Not one of the 6 analyzed mixed type tumors showed HER-2/neu expression. For the paired samples (preoperative biopsy samples and resected specimens) the concordance rate for HER-2/neu expression was 94.7%. CONCLUSIONS: According to high concordance rate in paired samples we consider it appropriate to evaluate HER2 expression on biopsy specimens, especially in unresectable cases, and to re-evaluate it on resected specimens if available, due to high heterogeneity of a gastric cancer.

Activation of cGAS-STING Pathway Is Associated with MSI-H Stage IV Colorectal Cancer.

Colorectal cancer is the second most common cause of cancer-related mortality in adults. Understanding colorectal tumorigenesis at both the cellular and molecular levels is crucial for developing effective treatment options. Forty-one biopsy samples from patients with metastatic CRC (mCRC) were collected at Split University Hospital in Croatia. A total of 41 patients (21 with microsatellite unstable tumours and 20 with microsatellite stable tumours) were randomly included in the study. Immunolabelling of cGAS and STING in metastatic CRC was performed and further complemented by histological classification, tumour grade, and KRAS, NRAS, and BRAF mutational status of mCRC. In bivariate analysis, elevated expression of cGAS and STING was positively associated with MSI-H colon cancer (Fisher's exact test, both p = 0.0203). Combined expression analysis of cGAS and STING showed a significantly higher percentage of patients with mCRC MSI-H with a fully or partially activated cGAS-STING signalling pathway (chi-square test, p = 0.0050). After adjusting for age, sex, and STING expression, increased cGAS expression remained significantly associated with MSI-H colon cancer in a multiple logistic regression model (ß = 1.588, SE = ±0.799, p = 0.047). The cGAS-STING signalling axis represents a compelling new target for optimization of immune checkpoint inhibitor therapeutic approaches in patients with MSI-H stage IV CRC.

Obstructive Giant Inflammatory Polyposis as a First Manifestation of Crohn's Disease: A Case Report.

Colonic inflammatory polyps (pseudopolyps) are common feature of inflammatory bowel diseases. They usually do not grow excessively, rarely reaching more than 15 mm in size, at which point they are termed giant inflammatory polyps. Clinical presentation of these polyps can vary greatly, ranging from being completely asymptomatic, usually detected incidentally, to abdominal cramps, rectal bleeding or intestinal obstruction. More importantly, giant inflammatory polyps can be easily mistaken for colonic malignancy, although without having malignant potential themselves. These polyps rarely regress with successful medical treatment of inflammatory bowel diseases and often require surgical treatment. We present an unusual case of giant inflammatory polyps which was the first presentation of inflammatory bowel disease. It was initially mistaken for colonic malignancy with intestinal obstruction, which led to surgical treatment.

DNA Methylome Changes of Muscle- and Neuronal-Related Processes Precede Bladder Cancer Invasiveness.

Bladder cancer (BC) is the ninth leading cause of cancer death with one of the highest recurrence rates among all cancers. One of the main risks for BC development is exposure to nitrosamines present in tobacco smoke or in other products. Aberrant epigenetic (DNA methylation) changes accompanied by deregulated gene expression are an important element of cancer pathogenesis. Therefore, we aimed to determine DNA methylation signatures and their impacts on gene expression in mice treated with N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN), a carcinogen similar to compounds found in tobacco smoke. Following BBN administration mice developed non-invasive or invasive bladder cancers. Surprisingly, muscle- and neuronal-related pathways emerged as the most affected in those tumors. Hypo- and hypermethylation changes were present within non-invasive BC, across CpGs mapping to the genes involved in muscle- and neuronal-related pathways, however, methylation differences were not sufficient to affect the expression of the majority of associated genes. Conversely, invasive tumors displayed hypermethylation changes that were linked with alterations in gene expression profiles. Together, these findings indicate that bladder cancer progression could be revealed through methylation profiling at the pre-invasive cancer stage that could assist monitoring of cancer patients and guide novel therapeutic approaches.